Recombinant adeno-associated virus (rAAV) vectors are one of the most promising in vivo gene delivery tools. Several features make rAAV vectors an ideal platform for gene transfer. However, the high homology with the parental wild-type virus, which often infects humans, poses limitations in terms of immune responses associated with this vector platform. Both humoral and cell-mediated immunity to wild-type AAV have been documented in healthy donors, and, at least in the case of anti-AAV antibodies, have been shown to have a potentially high impact on the outcome of gene transfer. While several factors can contribute to the overall immunogenicity of rAAV vectors, vector design and the total vector dose appear to be responsible of immune-mediated toxicities. While preclinical models have been less than ideal in predicting the outcome of gene transfer in humans, the current preclinical body of evidence clearly demonstrates that rAAV vectors can trigger both innate and adaptive immune responses. Data gathered from clinical trials offers key learnings on the immunogenicity of AAV vectors, highlighting challenges as well as the potential strategies that could help unlock the full therapeutic potential of in vivo gene transfer.

Low‐affinity MHC class I‐associated cryptic epitopes derived from self proteins overexpressed in a wide variety of human tumors or derived from antigens of viruses exhibiting a high mutation rate, could be interesting candidates for tumor and virus immunotherapy, respectively. However, identification of low‐affinity MHC‐associated epitopes comes up against their poor immunogenicity. Here we describe an approach that enhances immunogenicity of nonimmunogenic low‐affinity HLA‐A2.1‐binding peptides. It consists of modifying their sequence by introducing a tyrosine in the first position (P1Y). P1Y substitution enhances affinity of HLA‐A2.1‐associated peptides without altering their antigenic specificity. In fact, P1Y variants of ten nonimmunogenic low‐affinity peptides exhibited a 2.3‐ to 55‐fold higher binding affinity and/or stabilized the HLA‐A2.1 for at least 2 h more than the corresponding native peptides. More importantly, P1Y variants efficiently triggered in vivo native peptide‐specific CTL which also recognized the corresponding naturally processed epitope. The possibility for generating CTL against any low‐affinity HLA‐A2.1‐associated peptide provides us with the necessary tool for the identification of cryptic tumor and virus epitopes which could be used for peptide‐based immunotherapy.